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Increased amyloidogenic secretion in cerebellar granule cells undergoing apoptosis

By Cinzia Galli, Alessandra Piccini, Maria Teresa Ciotti, Loriana Castellani, Pietro Calissano, Damiano Zaccheo and Massimo Tabaton

Abstract

Some clues suggest that neuronal damage induces a secondary change of amyloid β protein (Aβ) metabolism. We investigated this possibility by analyzing the secretion of Aβ and processing of its precursor protein (amyloid precursor protein, APP) in an in vitro model of neuronal apoptosis. Primary cultures of rat cerebellar granule neurons were metabolically labeled with [35S]methionine. Apoptosis was induced by shifting extracellular KCl concentration from 25 mM to 5 mM for 6 h. Control and apoptotic neurons were then subjected to depolarization-stimulated secretion. Constitutive and stimulated secretion media and cell lysates were immunoprecipitated with antibodies recognizing regions of Aβ, full-length APP, α- and β-APP secreted forms. Immunoprecipitated proteins were separated by SDS/PAGE and quantitated with a PhosphorImager densitometer. Although intracellular full-length APP was not significantly changed after apoptosis, the monomeric and oligomeric forms of 4-kDa Aβ were 3-fold higher in depolarization-stimulated secretion compared with control neurons. Such increments were paralleled by a corresponding increase of the β-APPs/α-APPs ratio in apoptotic secretion. Immunofluorescence studies performed with an antibody recognizing an epitope located in the Aβ sequence showed that the Aβ signal observed in the cytoplasm and in the Golgi apparatus of control neurons is uniformly redistributed in the condensed cytoplasm of apoptotic cells. These studies indicate that neuronal apoptosis is associated with a significant increase of metabolic products derived from β-secretase cleavage and suggest that an overproduction of Aβ may be the consequence of neuronal damage from various causes

Topics: Biological Sciences
Publisher: The National Academy of Sciences
Year: 1998
OAI identifier: oai:pubmedcentral.nih.gov:18734
Provided by: PubMed Central
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